Phosphate (Pi) is the basic anionic component of the mineralized bone matrix, which is primarily hydroxyapatite. During bone resorption, the osteoclast is exposed to high concentrations of Pi released through demineralization of the bone matrix. Therefore, it is conceivable that the osteoclasts possess specific transport mechanisms for Pi uptake into the intracellular free Pi pool. Furthermore, Pi transport may be linked to maintenance of the ATP content of the osteoclast during the cyclical processes of migration, attachment and resorption. In this application, the principal investigator proposes that the activity of a NaPi-cotransporter plays a key role in maintaining substrate levels necessary for the energy requirements of the osteoclast. The applicant has evidence that the Na-dependent Pi cotransporter in the osteoclast localizes within a recycling endosomal pool which rapidly redistributes to the plasma membrane upon contact with bone. The goal of this proposal is to answer several questions which have arisen during the initial characterization of the Na-dependent phosphate (NaPi)-cotransporter in the osteoclast. These include: (1) what is the molecular identify of the NaPi-cotransporter in the osteoclast; (2) what is the nature of the recycling endosomal pool which contains the NaPi cotransporter; 3) what are the signals initiated by the attachment of osteoclasts to bone, leading to stimulation of Pi uptake; and (4) what is the contribution of the Na:Pi-cotransporter to normal osteoclast function. To address these issues, the Specific Aims of this application are: (1) to determine the molecular identify of the NaPi-cotransporter in the osteoclast; the applicant postulates that it is related, but not identical to the renal NaPi-2 isoform (rat, Type II family); (2) to determine the mechanisms of vesicular trafficking of the NaPi-cotransporter in the osteoclast. The applicant will focus on the initial signaling pathways which trigger the stimulatory responses and differential targeting of the NaPi-cotransporter; and (3) to determine the role of the NaPi-cotransporter in normal osteoclast function. The applicant proposes that phosphate transport is a process critical to normal osteoclast function. The efficacy of "knocking out" the function and expression of the NaPi-cotransporter will be assessed, both in vitro and in vivo. It is suggested by the applicant that the NaPi-cotransporter in the osteoclast may provide a potential target for novel therapeutic interventions in the treatment of osteoporosis.